Pulsed power and capillaries save time and improve results
Getting the best samples as efficiently as possible really matters, especially in processes such as single-cell sequencing (SCS). One area where this is important both in terms of efficiency and results is cancer research, in which scientists want to collect the most specific information as easily as possible. This often means finding appropriate technologies.
Scientists can apply SCS to improve the depth of information acquired in cancer research. To really understand this family of diseases, scientists want to collect the most specific information, but do so as easily as possible, which often means finding techniques that cut down on sample preparation, and efficiently handling samples really matters in SCS.
Nicholas Navin of The University of Texas MD Anderson Cancer Center (Houston), noted that SCS “is a powerful new tool for investigating evolution and diversity in cancer and understanding the role of rare cells in tumor progression,”1 adding, “The central problem inhibiting these studies is technical.”
Other experts echo this. Charles Gawad, St. Jude Children’s Research Hospital (Memphis, Tenn.) and his colleagues wrote: “[T]his field rests on the ability to study a single DNA molecule from individually isolated cells, a process that is technically challenging.”2 They explained, “Another critical component of obtaining genetic information from single cells is to amplify the single copy of a genome while minimizing the introduction of artifacts, such as amplification bias, genome loss, mutations and chimaeras.”
A time-saving sample preparation method could expand the use of SCS in many areas, including cancer research.
Faster fragments
Advanced Analytical Technologies (Ankeny, Iowa) developed its FEMTO Pulse Automated Pulsed-Field CE Instrument—winner of a 2016 R&D 100 Award—to separate and detect large, low-concentration DNA samples with the high degree of accuracy and precision necessary for SCS and other techniques. Steve Siembieda, vice president of commercialization at the company, says, “The FEMTO Pulse separates large DNA fragments 20 times faster than previous techniques and in an automated fashion, which saves a huge amount of time. Because it automates the runs and data analysis, hundreds of samples can be run every day with little effort.”
The platform’s high-voltage, pulsed-field power supply drives the separation of nucleic acids in capillaries. Along with an optimized detection system and a sensitive intercalating dye, this creates a fast and sensitive process. “Other technologies detect picograms of nucleic acids, and the FEMTO Pulse—as the name suggests—detects in the femtograms, which allows analysis of a single cell’s worth of nucleic acid,” Siembieda says. “That sensitivity allows people to save on the amount of sample that they need.”
In SCS, scientists need to save all of the sample that they can. Some will be used for quality control to make sure the sample includes nucleic acids before going to the trouble and expense of sequencing it. “With the FEMTO Pulse,” says Siembieda, “researchers can use a small portion of captured nucleic acids for QC and leave the rest for analysis.”
Sidestepping amplification
Many studies of nucleic acids increase the quantity of the sample with the polymerase chain reaction (PCR). The sensitivity of the FEMTO Pulse can make PCR unnecessary in some cases or reduce it dramatically in others. “With such sensitive detection and using an intercalation dye, you may be able to find virus in a sample without PCR,” Siembieda explains. “Or researchers may amplify their isolated DNA with fewer PCR cycles to reduce amplification bias.”
Some methods of working with nucleic acids also require knowing something about the sequence ahead of time, but not so with this technology. “Our intercalating dye will stain whatever nucleic acid is there,” says Siembieda, “and we don’t need to know anything about the DNA.” That means the platform can also be used to analyze unknowns.
For SCS and other methods that work with small amounts of nucleic acids, the right sample preparation can save time and still provide high-end outcomes.
References
- Navin, N.E. The first five years of single-cell cancer genomics and beyond. Genome Res. 2015; doi: 10.1101/gr.191098.115.
- Gawad, C.; Winston, K.; Quake, S.R.. Single-cell genome sequencing: current state of the science. Nat. Rev. Genetics 2016; doi: 10.1038/nrg.2015.16.